Enterococci rank among most common causes of hospital-acquired infection, making them a leading public health concern. Many of the enterococci that cause these infections are resistant to multiple antibiotics - and with increasing frequency, to all antibiotics. During the 1980's one genetic lineage of E. faecalis emerged that has acquired resistances to all antibiotics and is highly virulent. We and others found members of this lineage to be passed from patient to patient within hospital wards, and from hospital to hospital. Over a quarter of the genome of this E. faecalis lineage consists of acquired mobile DMA.We propose a series of experiments to follow up our description of a pathogenicity island in this virulent, antibiotic resistant lineage of E. faecalis. Initial studies examine the evolution of the PAI in strains collected since the early days of the 20th Century, and are designed to identify its pattern of inheritance and the accretion of traits. The proposed studies then ask about the stability of the element and its rate and mechanism of transfer. In the process, isogenic strains lacking the PAI and plasmids that occur natively will be generated, which then allows comparison of the ability of these strains to acquire new traits to that of other E. faecalis lineages. This tests the hypothesis that the PAI, as well as unlinked genes for vancomycin resistance and b-lactamase production, were independently discovered in one particular E. faecalis lineage because this lineage is prolific at acquiring and exchanging mobile elements. Finally, the availability of isogenic strains permits us to ask whether the PAI confers an advantage either in growth in a complex human Gl tract-derived consortium, or in colonization using a novel Drosophila model.